Most, if not all, cellular processes are regulated by timely degradation of regulatory proteins through the ubiquitin system. Specific substrate proteins are targeted for degradation through the activity of E3 ubiquitin ligases. We previously discovered that two small RING finger proteins, ROC1 and ROC2, function as the essential catalytic components of the largest family of E3 ligase complexes, the cullin-RING E3 ligases (CRLs). We also discovered that cullin 3 and cullin 4 could assemble as many as 200 and 100 distinct CRL3 and CRL4 E3 complexes, respectively. In this application, we propose two lines of research aimed at understanding the function and mechanism of two specific CRL4 E3 ubiquitin ligase complexes, CRL4G2, and CRL4WDR5, and how alterations of these CRL4 complexes contribute to two major human diseases, heart failure and X-linked mental retardation. In the first Aim, we will examine the molecular function of CRL4 G2 and its role in cardiovascular disease. We present data demonstrating that a WD40 protein, G-protein subunit 2 (G2), associates with DDB1- CUL4A to target G protein-coupled receptor kinases (GRK2), for ubiquitylation. Elevated GRK2 has previously been associated with myocardial infarction, heart failure, portal hypertension, insulin resistance, and Alzheimer's disease. We propose first to explore how broadly CRL4 is involved in the regulation of G- protein coupled receptor (GPCR) signaling by determining the interaction of CRL4 with other G and GRK proteins. We will then focus on one well-characterized GPCR signaling pathway, -adrenergic receptors ( - AR) signaling, to determine the mechanism of CRL4G2 mediated GRK2 ubiquitylation. We will also determine whether loss of Cul4a impairs heart function in mouse. In the second Aim, we present data demonstrating that CUL4B, one of the most frequently mutated genes in X-linked mental retardation (XLMR) patients, but not CUL4A, is a nuclear E3 ligase of WDR5, a WD40 protein and an essential component for histone H3K4 trimethylation. We propose to determine the function of CUL4B-mediated WDR5 ubiquitylation in neuronal cell proliferation, survival and differentiation. We also propose to develop and characterize a novel mouse model, conditional Cul4b mutant mouse strain, to determine the function of Cul4b in brain. Finally, we propose to determine broadly the function of CUL4B in the regulation of chromatin modification and gene expression.